Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays

Detalhes bibliográficos
Autor(a) principal: Manta, Fernanda Saloum de Neves
Data de Publicação: 2020
Outros Autores: Leal-Calvo, Thyago, Moreira, Suelen Justo M., Marques, Brunna L. C., Ribeiro-Alves, Marcelo, Rosa, Patrícia S., Nery, José Augusto C., Rampazzo, Rita de Cássia Pontello, Costa, Alexandre Dias Tavares, Krieger, Marco Aurelio, Moraes, Milton Ozório
Tipo de documento: Artigo
Idioma: por
Título da fonte: Repositório Institucional da FIOCRUZ (ARCA)
Texto Completo: https://www.arca.fiocruz.br/handle/icict/42019
Resumo: Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.
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spelling Manta, Fernanda Saloum de NevesLeal-Calvo, ThyagoMoreira, Suelen Justo M.Marques, Brunna L. C.Ribeiro-Alves, MarceloRosa, Patrícia S.Nery, José Augusto C.Rampazzo, Rita de Cássia PontelloCosta, Alexandre Dias TavaresKrieger, Marco AurelioMoraes, Milton Ozório2020-06-30T20:54:22Z2020-06-30T20:54:22Z2020MANTA, Fernanda Saloum de Neves et al. Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays. PLoS Neglected Tropical Diseases, v. 14, n. 5, p. 1-15, 2020.1935-2735https://www.arca.fiocruz.br/handle/icict/4201910.1371/journal.pntd.00083251935-2735porPublic Library of SciencePCRLeprosyPolymerase Chain ReactionReagent Kits, DiagnosticLepraReacción en Cadena de la PolimerasaJuego de Reactivos para DiagnósticoHanseníaseReação em Cadeia da PolimeraseKit de Reagentes para DiagnósticoUltra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assaysinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleFundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Nacional de Infectologia Evandro Chagas. Laboratório de Pesquisa Clínica em DST-AIDS. Rio de Janeiro, RJ, Brasil.Instituto Lauro de Souza Lima. Bauru, SP, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto de Biologia Molecular do Paraná. Curitiba, PR, Brasil.Fundação Oswaldo Cruz. Instituto de Biologia Molecular do Paraná. Curitiba, PR, Brasil. / Fundação Oswaldo Cruz. Instituto Carlos Chagas. Curitiba, PR, Brasil.Fundação Oswaldo Cruz. Instituto de Biologia Molecular do Paraná. Curitiba, PR, Brasil. / Fundação Oswaldo Cruz. Instituto Carlos Chagas. Curitiba, PR, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Leprosy urgently needs a precise and early diagnostic tool. The sensitivity of the direct (bacilli staining, Mycobacterium leprae DNA) and indirect (antibody levels, T cell assays) diagnostics methods vary based on the clinical form. Recently, PCR-based M. leprae DNA detection has been shown to differentially diagnose leprosy from other dermatological conditions. However, accuracy can still be improved, especially for use with less invasive clinical samples. We tested different commercial DNA extraction kits: DNeasy Blood & Tissue, QIAamp DNA Microbiome, Maxwell 16 DNA Purification, PowerSoil DNA Isolation; as well as in-house phenol-chloroform and Trizol/FastPrep methods. Extraction was performed on M. leprae-infected mouse footpads and different clinical samples of leprosy patients (skin biopsies and scrapings, lesion, oral and nasal swabs, body hair, blood on FTA cards, peripheral whole blood). We observed that the Microbiome kit was able to enrich for mycobacterial DNA, most likely due the enzymatic digestion cocktail along with mechanical disruption involved in this method. Consequently, we had a significant increase in sensitivity in skin biopsies from paucibacillary leprosy patients using a duplex qPCR targeting 16S rRNA (M. leprae) and 18S rRNA (mammal) in the StepOnePlus system. Our data showed that the presence of M. leprae DNA was best detected in skin biopsies and skin scrapings, independent of the extraction method or the clinical form. For multibacillary patients, detection of M. leprae DNA in nasal swabs indicates the possibility of having a much less invasive sample that can be used for the purposes of DNA sequencing for relapse analysis and drug resistance monitoring. Overall, DNA extracted with the Microbiome kit presented the best bacilli detection rate for paucibacillary cases, indicating that investments in extraction methods with mechanical and DNA digestion should be made.info:eu-repo/semantics/openAccessreponame:Repositório Institucional da FIOCRUZ (ARCA)instname:Fundação Oswaldo Cruz (FIOCRUZ)instacron:FIOCRUZLICENSElicense.txtlicense.txttext/plain; charset=utf-83084https://www.arca.fiocruz.br/bitstream/icict/42019/1/license.txt783568c2893d2e25a99990b126be1772MD51ORIGINALjournal.pntd.000832.pdfjournal.pntd.000832.pdfapplication/pdf1434923https://www.arca.fiocruz.br/bitstream/icict/42019/2/journal.pntd.000832.pdf8cf1cbd5c7b0c3c228cb3c2ed1cb4611MD52TEXTjournal.pntd.000832.pdf.txtjournal.pntd.000832.pdf.txtExtracted texttext/plain49711https://www.arca.fiocruz.br/bitstream/icict/42019/3/journal.pntd.000832.pdf.txt790c185fc532fb6c98dcf0ec020e1680MD53icict/420192023-08-02 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dc.title.pt_BR.fl_str_mv Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays
title Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays
spellingShingle Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays
Manta, Fernanda Saloum de Neves
PCR
Leprosy
Polymerase Chain Reaction
Reagent Kits, Diagnostic
Lepra
Reacción en Cadena de la Polimerasa
Juego de Reactivos para Diagnóstico
Hanseníase
Reação em Cadeia da Polimerase
Kit de Reagentes para Diagnóstico
title_short Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays
title_full Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays
title_fullStr Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays
title_full_unstemmed Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays
title_sort Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays
author Manta, Fernanda Saloum de Neves
author_facet Manta, Fernanda Saloum de Neves
Leal-Calvo, Thyago
Moreira, Suelen Justo M.
Marques, Brunna L. C.
Ribeiro-Alves, Marcelo
Rosa, Patrícia S.
Nery, José Augusto C.
Rampazzo, Rita de Cássia Pontello
Costa, Alexandre Dias Tavares
Krieger, Marco Aurelio
Moraes, Milton Ozório
author_role author
author2 Leal-Calvo, Thyago
Moreira, Suelen Justo M.
Marques, Brunna L. C.
Ribeiro-Alves, Marcelo
Rosa, Patrícia S.
Nery, José Augusto C.
Rampazzo, Rita de Cássia Pontello
Costa, Alexandre Dias Tavares
Krieger, Marco Aurelio
Moraes, Milton Ozório
author2_role author
author
author
author
author
author
author
author
author
author
dc.contributor.author.fl_str_mv Manta, Fernanda Saloum de Neves
Leal-Calvo, Thyago
Moreira, Suelen Justo M.
Marques, Brunna L. C.
Ribeiro-Alves, Marcelo
Rosa, Patrícia S.
Nery, José Augusto C.
Rampazzo, Rita de Cássia Pontello
Costa, Alexandre Dias Tavares
Krieger, Marco Aurelio
Moraes, Milton Ozório
dc.subject.other.pt_BR.fl_str_mv PCR
topic PCR
Leprosy
Polymerase Chain Reaction
Reagent Kits, Diagnostic
Lepra
Reacción en Cadena de la Polimerasa
Juego de Reactivos para Diagnóstico
Hanseníase
Reação em Cadeia da Polimerase
Kit de Reagentes para Diagnóstico
dc.subject.en.pt_BR.fl_str_mv Leprosy
Polymerase Chain Reaction
Reagent Kits, Diagnostic
dc.subject.es.pt_BR.fl_str_mv Lepra
Reacción en Cadena de la Polimerasa
Juego de Reactivos para Diagnóstico
dc.subject.decs.pt_BR.fl_str_mv Hanseníase
Reação em Cadeia da Polimerase
Kit de Reagentes para Diagnóstico
description Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.
publishDate 2020
dc.date.accessioned.fl_str_mv 2020-06-30T20:54:22Z
dc.date.available.fl_str_mv 2020-06-30T20:54:22Z
dc.date.issued.fl_str_mv 2020
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
dc.type.driver.fl_str_mv info:eu-repo/semantics/article
format article
status_str publishedVersion
dc.identifier.citation.fl_str_mv MANTA, Fernanda Saloum de Neves et al. Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays. PLoS Neglected Tropical Diseases, v. 14, n. 5, p. 1-15, 2020.
dc.identifier.uri.fl_str_mv https://www.arca.fiocruz.br/handle/icict/42019
dc.identifier.issn.pt_BR.fl_str_mv 1935-2735
dc.identifier.doi.none.fl_str_mv 10.1371/journal.pntd.0008325
dc.identifier.eissn.none.fl_str_mv 1935-2735
identifier_str_mv MANTA, Fernanda Saloum de Neves et al. Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays. PLoS Neglected Tropical Diseases, v. 14, n. 5, p. 1-15, 2020.
1935-2735
10.1371/journal.pntd.0008325
url https://www.arca.fiocruz.br/handle/icict/42019
dc.language.iso.fl_str_mv por
language por
dc.rights.driver.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.publisher.none.fl_str_mv Public Library of Science
publisher.none.fl_str_mv Public Library of Science
dc.source.none.fl_str_mv reponame:Repositório Institucional da FIOCRUZ (ARCA)
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collection Repositório Institucional da FIOCRUZ (ARCA)
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